Objective:To analyze and identify the expression profiles of oxidative stress-related genes and circular RNAs(circRNAs)in ricin toxin(RT)-induced pyroptosis of mouse mononuclear macrophages(RAW264.7)using transcriptome sequencing and bioinformatics technology,and to preliminarily analyze their potential functions.Methods:The macrophages(RAW264.7 cells)were treated with RT to establish a cell pyroptosis model and divided into control group,40 μg·L-1 RT group,and 80 ng/mL RT group.Transmission electron microscope(TEM)was used to observe the morphology of the RAW264.7 cells in various groups;Western blotting method was used to detect the expression levels of the pyroptosis pathway-related proteins in the cells in various groups;80 μg·L-1 RT was selected for subsequent experiments.Transcriptome sequencing(RNA-Seq)was performed to obtain the circRNA and mRNA expression profiles in the RAW264.7 cells in control group and RT-treated group,followed by bioinformatics analysis.Results:Compared with control group,the cells in 40 and 80 μg·L-1 RT groups underwent morphological changes;the cells in RT groups showed obvious pyroptosis-like morphological changes,characterized by significant swelling of dying cells and the appearance of characteristic large bubbles on the plasma membrane.Compared with control group,the expression level of gasdermin DN-terminal fragment(GSDMD-N)protein in 40 and 80 μg·L-1 RT groups was increased(P<0.05);compared with 40 μg·L-1 RT group,the expression level of GSDMD-N protein in the cells in 80 μg·L-1 RT group was increased(P<0.05);therefore,the subsequent experiments used the RT concentration of 80 μg·L-1.A total of 2930 differentially expressed messenger RNAs(mRNAs)and 24 differentially expressed circRNAs were identified.The constructed circRNA-microRNA(miRNA)-mRNA competing endogenous RNA(ceRNA)regulatory network consisted of 7 circRNAs,12 miRNAs and 13 mRNAs.Gene Ontology(GO)functional enrichment analysis showed that in biological process(BP),the functions regulated by differentially expressed genes mainly included immune response and oxidative stress response;in cellular component(CC),differentially expressed genes were mainly localized to the external side of plasma membrane and cell leading edge;in molecular function(MF),they were mainly involved in transporter transmembrane activity and hormone receptor binding.Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analysis showed that differentially expressed genes were mainly enriched in Toll-like receptor signaling pathway,Forkhead box O(FoxO)signaling pathway and nuclear factor kappa-light-chain-enhancer of activated B cells(NF-κB)signaling pathway.In the protein-protein interaction(PPI)network,the top 10 hub genes with the highest connectivity were screened by CytoHubba,including matrix metalloproteinase 9(MMP9),superoxide dismutase 2(SOD2),and v-src sarcoma viral oncogene homolog(Src).Conclusion:The expression profiles of oxidative stress-related genes and circRNAs in RAW264.7 cells are altered after RT treatment.The screened differentially expressed circRNAs and mRNAs may serve as potential targets to regulate RT-induced pyroptosis in RAW264.7 cells through oxidative stress pathways.

Objective:To discuss the damage effect of vesicular stomatitis virus(VSV)on the vascular endothelial(VE)barrier,and to clarify its mechanism.Methods:The canine kidney cells were used to amplify VSV.The half tissue culture infective dose(TCID50)of VSV was determined using mouse brain endothelial tumor bEnd.3 cells,and subsequent experiment was conducted using 300 times the TCID50.The bEnd.3 cells were divided into infection 0 h group,infection 4 h group,infection 8 h group,and infection 12 h group for VE barrier damage experiments due to VSV infection.The bEnd.3 cells were also divided into control group,infection group,and correction group for experiments to inhibit the VSV replication and restore the VE barrier.The bEnd.3 cells were inoculated into Transwell chambers to construct an in vitro VE barrier model.Cell voltage resistance meter was used to detect the transepithelial resistance(TER)in various groups after the bEnd.3 cells were infected with VSV at different time points;fluorescein isothiocyanate-dextran leakage assay was used to detect the permeability coefficients of the cells in various groups;immunofluorescence staining was used to observe the localization changes of VE-cadherin,β-catenin,and phosphorylated β-catenin(p-β-catenin)in cytoskeleton and adherens junctions(AJs)of the bEnd.3 cells after VSV infection;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of Wnt and β-catenin mRNA in the cells in various groups;Western blotting method was used to detect the expression levels of Wnt,β-catenin,and p-β-catenin proteins in the cells in various groups.Results:The TCID50 of VSV was 10-4.5·100 μL-1.TheTranswell chamber experiment results showed that compared with infection 0 h group,the TERs in the cells in the other groups were significantly decreased(P＜0.05),and the permeability coefficients were significantly increased(P＜0.05).The immunofluorescence staining results showed that compared with control group,the cytoskeleton of the bEnd.3 cells in infection group was disordered,the cell gaps was increased,the linear index of AJs was significantly decreased(P＜0.05),and β-catenin and p-β-catenin translocated from the cell membrane to the perinuclear area.The RT-qPCR results showed that compared with infection 0 h group,the expression levels of Wnt mRNA in the cells in the other groups were significantly decreased(P＜0.05),while the expression levels of β-catenin mRNA showed no statistically significant difference(P＞0.05).The Western blotting results showed that compared with infection 0 h group,the expression levels of Wnt protein in the cells in the other groups were significantly decreased(P＜0.05),the expression levels of β-catenin showed no statistically significant differences(P＞0.05),and the expression levels of p-β-catenin were significantly increased(P＜0.05).After inhibiting the VSV replication and correcting the low density lipoprotein receptor(LDLR)abnormalities,the Transwell chamber experiment results showed that compared with infection group,the TER in the cells in correction group was significantly increased(P＜0.05),and the permeability coefficient was significantly decreased(P＜0.05).The immunofluorescence staining results showed that compared with infection group,the gaps in the cells in correction group were reduced,and the perinuclear aggregation of β-catenin and p-β-catenin in the cells was restrained.The RT-qPCR results showed that compared with infection group,the expression level of Wnt mRNA in the cells in correction group was significantly increased(P＜0.05).The Western blotting results showed that compared with infection group,the expression level of Wnt protein in the cells in correction group was significantly increased(P＜0.05),the expression level of β-catenin showed no statistically significant difference(P＞0.05),and the expression level of p-P-catenin was significantly decreased(P＜0.05).Conclusion:VSV infection can cause the LDLR inactivation,reduce the expression level of Wnt protein,increase the phosphorylation level of β-catenin and cause its internalization,disrupt the stability of AJs,and ultimately lead to VE barrier damage.

Gradual distraction with an external fixator is a widely used treatment for severe postburn ankle contracture (SPAC). However, application of external fixators is complex, and conventional two-dimensional (2D) imaging-based surgical planning is not particularly helpful due to a lack of spatial geometry. The purpose of this study was to evaluate the surgical planning process for this procedure with patient-specific three-dimension-printed models (3DPMs). In this study, patients coming from two centers were divided into two cohorts (3DPM group vs. control group) depending on whether a 3DPM was used for preoperative surgical planning. Operation duration, improvement in metatarsal-tibial angle (MTA), range of motion (ROM), the American Orthopedic Foot and Ankle Society (AOFAS) scores, complications, and patient-reported satisfaction were compared between two groups. The 3DPM group had significantly shorter operation duration than the control group ((2.0±0.3) h vs. (3.2±0.3) h,

Objective To explore a novel strategy to repair traumatic soft tissue defects in the head and face by tissue expansion in the early stage.Methods Eighteen patients with traumatic soft tissue defects were treated with thorough debridement leaving the wound unclosed or simply closed with thin split-thickness scalp skin grafting,and adjacent expander implantation in the early stage and expanded flap transposition in the second stage.Results There were 11 male patients and 7 female patients ranging in age from 3.5 to 40 years [mean,(19.4±12.2) years],with average 15 months follow-up (range,3-67 months).The average expansion time was 74.3 days (range,53-96 days).The total of 18 patients with 22 expanders were treated with satisfactory results.All the flaps survived and the skin color,texture and contour well matched to those of the peripheral tissue.Only one complication of infection happened in the 18 cases (5.56％) and total 22 expanders (4.55％),which was similar to the rate reported in the literature.There were no any other complications related to the expander.Conclusions Debridement and tissue expansion in the early stage have been proved a more effective strategy to repair traumatic soft tissue defects in the head and face,which can not only achieve satisfactory color,unbulky and well matched texture similar to normal,but also can avoid unnecessary donor site injuries and does not increase the infection rate of tissue expansion.

Objective To investigate the expression and effect of miR-22-3p in non-small cell lung cancer (NSCLC). Methods The miR-22-3p expression level in seventy-six NSCLC tissues and para-cancer tissues was detected by qRT-PCR. The relationship between the expression of miR-22-3p and gender,age,tumor size,histolo-gy grade,pathological type and lymph node metastasis was analyzed. The function of miR-22-3p on the prolifera-tion of NSCLC cells was tested by growth curve assay. Target genes of miR-22-3p were predicted by online software Targetscan. Luciferase reporter assay and qRT-PCR was used to certificate the prediction. Results The expression of miR-22-3p was increased in NSCLC tissues than the para-cancer tissues and was correlated to lymph node metas-tasis. Overexpression of miR-22-3p could suppress the proliferation of A549 cells. Astrocyte-Elevated Gene-1(AEG-1) was predicted to be a target of miR-22-3p. MiR-22-3p was revealed to bind to AEG-13′UTR by luciferase report-er assay. Overexpression of miR-22-3p could inhibit the expression of AEG-1 in A549 cells. Suppression of miR-22-3p could increase AEG-1 expression. Conclusion MiR-22-3p could inhibit the proliferation of NSCLC by tar-geting AEG-1.

Objective To investigate the therapeutic effect of ambroxol hydrochloride on severe pneumonia, and the influence on the inflammatory cytokines of TNF-alpha and hs-CRP etc.MethodsThe clinical data of 70 patients with severe pneumonia patients treated in our hospital from December 2014 to August 2016 were retrospectively analyzed.The patients were divided into control group(n=30) and observation group(n=40) according to the treatment methods, the control group was given routine treatment, the observation group was given ambroxol hydrochloride on the basis of conventional treatment.The therapeutic effect of the two groups was observed, the differences of serum levels of inflammatory cytokines, pulmonary function, cough and expectoration symptom scores of the two groups were compared before and after treatment.ResultsThe effective rate of the observation group was 97.50%, which was significantly higher than that of the control group(80.00%);There was no significant difference in serum inflammatory cytokine levels between the two groups before treatment, after treatment,the levels of IL-6, IL-8, hs-CRP and TNF-alpha the observation group were lower than those in the control group;There was no significant difference in the indexes of lung function between the two groups before treatment.After treatment, the levels of FVC, FEV1 and FEV1/FVC in the observation group were higher than those in the control group;There was no significant difference in symptom score between the two groups before treatment.After treatment, cough, sputum volume, cough ease and wheezing scores in the observation group were lower than in the control group.ConclusionAmbroxol hydrochloride has a good therapeutic effect on severe pneumonia,which can significantly reduce the levels of inflammatory cytokines such as TNF-α and hs-CRP, and improve the related symptoms, and has good application value.

Objective To observe the effect of tiotropium combined Seretide in the treatment of elderly patients with moderate to severe COPD clinical curative effect and the influence on the quality of life of patients.Methods114 cases of moderate or severe COPD in our hospital from January 2014 to January 2016,were randomly divided into observation group and control group, 57 cases in each group.The control group was treated with Seretide treatment, the observation group were treated by tiotropium combined with Seretide treatment, compared two groups of patients with curative effect and quality of life.ResultsBefore treatment, there were no significant differences in lung function index, MRC score, 6MWT and WHOQOL-100 scores between the two groups;After treatment, FEV1/FVC and FEV1% in the observation group were significantly higher than in the control group, and the difference was statistically significant (P<0.05);the dyspnea severity score (MRC) and six minute walk test (6MWT) in the observation group were significantly better than those in the control group, and the differences were statistically significant (P<0.05);the quality of life score (WHOQOL-100) in the observation group was significantly higher than in the control group, the difference was statistically significant (P<0.05).ConclusionTiotropium bromide combined with nimesulide in the treatment of elderly patients with moderate or severe COPD is effective, and can effectively improve the level of pulmonary function, improve the quality of life of patients.

Objective To explore the significance of detecting miR-145 hypermethylation in non-small cell lung cancer (NSCLC). Methods Seventy-five NSCLC tissues and adjacent tissues was collected from May 2012 to January 2015. The methylation status of miR-145 promoter was tested by bisulfite sequencing PCR (BSP) and the expression of miR-145 evaluated by ISH. The correlation between miR-145 methylation and clinical parameters , and the relationship between miR-145 methylation and expression were statistically analyzed. Results The hypermethy-lation ratio of miR-145 in NSCLC tissues was higher than that in the adjacent tissues. Hypermethylation of miR-145 was correlated with NSCLC differentiation stage, but not with gender and age. In NSCLC, miR-145 hypermethyla-tion was negatively related with its expression. Conclusion MiR-145 hypermethylation is closely related with dif-ferentiation stage in NSCLC tissues. MiR-145 hypermethylation may be a potential biomarker for estimating NSCLC differentiation stage.

Objective To study the increased radiosensitivity of tumor cells by triterpene acids of loquat leaf (TAL) and mechanism.Methods C6 and MG63 cells were pretreated by TAL,and then were exposed to X-rays at 1,2,3,5,7 Gy.Clonogenic assay was used to evaluate those tumor cells survival fraction (SF) to calculate the sensitization enhancement ratios (SER) of TAL.The cellular micronuclei ratios of tumor cells were analyzed by micronuclei assay.Additionally,the changes of Rad51 and XRCC4 levels were observed by RT-PCR and Western blot.Results D0 values were decreased to 1.31 and 2.85 Gy in C6 and MG63 cells by TAL pretreatment respectively.The SER value of the effect of TAL on C6 and MG63 cells was 1.73 and 2.04,respectively.There was a statistical difference in the cellular micronuclei ratios between tumor cells with TAL above 2 Gy and those without TAL (C6:t =-8.372--2.476,P ＜0.05 ; MG63:t =-4.03--2.557,P ＜ 0.05).TAL attenuated the expression of XRCC4 at transcriptional and translational level,but not for Rad51,a key gene in homologous recombination repair (HRR).Conclusions TAL pretreatment could increase the lethal effect of X-rays on tumor cells in vitro.The mechanism might be involved in the inhibition of non-homologous end joining (NHEJ).

Objective To explore a better method to repair cervical cicatricle contracture deformity.Methods The cervicai cicatrical contraction deformity was repaird by the cervico-shoulder island skin flap with blood supply crossing and abundant anastomotic branches of the neck-shoulder blood vessels. After removing the cervical scar, the flaps were designed according to the size of the wound to be repaired, and the clinical anatomy of crossing and abundant anastomotic branches of the neck-shoulder blood vessels. The secondary wound of donor site was closed directly if the donor region was small, or closed by skin graft. Results Sixteen patients in this clinic were operated with this method, and venous return dysfunction occurred in 1 case, but blood circulation was improved after treatment. All the flaps survived with good appearance and texture. The extensive flap was created and transferred to the anterior neck by one stage without pre-expansion which met some patients'requirement of repairing cervical cicatricle contracture deformity. Conclusion The cervico-shoulder island skin flap, with abundant blood supply, is a new method to repair cervical cicatricle contracture deformity.